Biological processes, specifically processes involving cells maintained in a fluid environment, are of increasing importance, because they are the means by which the unique synthetic and catalytic capabilities of prokaryotic and eukaryotic cells are put to work in manufacturing and other industrial processes. Examples of such processes include the production of proteins, antibodies, and other molecules useful, for example, in pharmaceutical applications and waste treatment processes, and industrial products, such as, for example, materials produced for foods, chemical processing, and agriculture, etc. Examples of such industrial products include enzymes for the textile, food, pulp and paper industries; feedstocks for chemical processes, such as acetone, butanol, ethanol, and acetic acid; biopolymers such as guar and carrageenan in foods and degradable packaging; dyes; flavors and fragrances; bioadhesives; and bioadsorbants and supports, for example, chitin.
Biological cells must be surrounded by water, and require nutrients (e.g., carbon, hydrogen, phosphorus, potassium, nitrogen, sulfur, calcium, iron, magnesium, manganese, boron, copper, and others), electron acceptors (oxygen for aerobes, e.g., nitrate or sulfate for anaerobes) and electron donors (e.g., carbohydrates). "Bioreactors" that contain and support bioprocesses, are designed to maintain appropriate process conditions, such as temperature, as well as nutrient supply and waste removal. Traditionally, biological cells have been submerged in vats or tanks of aqueous nutrient medium. However, such bioreactors are limited in their ability to efficiently facilitate the mass transport of nutrients and waste products, particularly gases, to and from biological cells. The delivery of sufficient oxygen from outside the bioreactor to meet the high oxygen demand of some processes is particularly problematic in such submerged tank reactors.
One common type of submerged tank bioreactor is the continuous stirred tank reactor (CSTR). The CSTR is essentially a closed, cylindrical vat with a vertical shaft projecting through the top or bottom with paddles attached. Rotation of the shaft stirs the bioreactor contents (aqueous medium and cells) and accomplishes mass transport through mechanical agitation. Cells are suspended in aqueous medium, and oxygen is delivered to the cells by a combination of gas injection and stirring by the revolving paddles.
There is significant potential for medium contamination in the CSTR, however. The seals for the rotating shaft are the primary route for the introduction of external contaminants into axenic cultures. In addition, control of heat transfer (for example, cooling fermentation broths which overheat due to the heat of metabolism) can be difficult. Oxygen delivery to the cells is also generally inefficient, as noted above. In order to transport sufficient oxygen to aerobic cells, the medium must be vigorously stirred. The resulting shear forces generated by the revolving paddles cause cell trauma, senescence, and often death. The dilemma is that many cells, particularly mammalian cells, which are sensitive to shear and which require high levels of oxygen, will die in high-shear environments; however, the same cells will die of anoxia if shear forces are reduced to tolerable levels. Also, the high energy input required for gas injection, mechanical stirring, and cooling of a CSTR is expensive.
Various bioprocess systems and reactor designs have been developed to solve this problem, including fixed film reactors and bubble columns. In fixed film reactors, cells are fixed to a surface and are stationary while the medium moves past them. In bubble columns, downward falling cells are suspended in medium by upward moving gas bubbles, and the bubble movement results in mass transfer of the gas in the bubbles. In these systems, shear is lower than in CSTRs, but the surface area of contact between the cells and the gas phase is a rate limiting factor.
In order to overcome the limitations of these processes, a number of methods for carrying out biological processes in foams have been developed. These processes employ foams that are essentially reticulated networks of gas/fluid interfaces of high surface area as the culture medium for cells, which are present in the liquid thin film with the reactants of a desired bioprocess reaction. Because the cells are proximal to the gas interface, improved mass transport rates and, therefore, faster process rates, with less energy input, can be achieved.
U.S. Pat. No. 3,677,895 to Hashimoto ("Hashimoto") describes a process for reacting paraffinic hydrocarbons with microbial cells to produce protein in a CSTR. The culture medium is an aqueous foam comprising a discontinuous gas phase with oxygen or a gaseous hydrocarbon and a continuous aqueous interface surrounding the gaseous reactant with nutrients, microorganisms, and foam stabilizers to maintain the foamed state. Hashimoto discloses batch, CSTR type reactors, and a centrifuge is used following fermentation to separate the microorganisms from the medium.
U.S. Pat. Nos. Reissue 30,543 and U.S. Pat. No. 4,340,677 to Hitzman, et al. ("Hitzman I") and Hitzman ("Hitzman II"), respectively, each relate to fermentation processes in a foam. The reactors disclosed are essentially batch reactors, although Hitzman I describes carrying out both "batch" and "continuous" fermentation processes. In the continuous process, oxygen or air, nutrient medium, and alcohol are continuously introduced into the reactor throughout the process run. Hitzman I and II each employ mechanical foam breakers.
Pumped loop reactors for carrying out bioprocesses utilizing foams are described in U.S. Pat. No. 5,021,069 to Whellock, et al. ("Whellock"). Whellock is primarily concerned with bioleaching processes, where the relatively high turbulence and shear conditions maintained in the reactor columns are desirable.
High shear forces are present, however, in the foam bioprocesses of Hashimoto, Hitzman I and Hitzman II, as well. The production of foam in cell-containing medium by mechanical stirring and/or rapid injection of gas exerts potentially harmful shear forces on delicate cells, as does the breaking of foam by with, for example, rotating surfaces as in Hitzman II. Cell trauma and death resulting from these forces limit process rates and yields, and increase the cost of many processes. The present invention is directed to overcoming the deficiencies in the art.